Functional Characterization of Glycosyltransferases from Saussurea involucrata Involved in Rutin Biosynthesis
4 affiliations
- 1Ministry of Education Key Laboratory of Chinese Medicinal Resource from Lingnan, School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou 510006, China
- 2Research Center of Chinese Herbal Resource Science and Engineering, Guangzhou University of Chinese Medicine, Guangzhou 510006, China
- 3Dongguan Institute, Guangzhou University of Chinese Medicine, Dongguan 523808, China
- 4Chinese Medicine Guangdong Laboratory, Hengqin, Zhuhai 519031, China
Abstract
Saussurea involucrata is a medicinal plant rich in flavonoids, but the glycosyltransferases (GTs) responsible for flavonoid glycosylation in this species remain largely uncharacterized. Here, five GT genes (SiUGT03, SiUGT09, SiUGT60, SiRhaT68-1, and SiRhaT68-2) were identified. Four of the five proteins (SiUGT03, SiUGT09, SiUGT60, and SiRhaT68-1) possessed an intact PSPG-box, and phylogenetic analysis placed SiUGT03 in the 3GT clade and SiRhaT68-1 in the 1,6GT clade. Recombinant proteins were expressed in Escherichia coli as soluble maltose-binding protein (MBP) fusions and purified. In vitro enzymatic assays showed that SiUGT03 catalyzed the 3-O-glucosylation of quercetin to isoquercetin (quercetin-3-O-glucoside), whereas SiRhaT68-1 catalyzed the 1,6-rhamnosylation of isoquercetin to rutin. Among the substrates tested, SiUGT03 was active only toward the flavonols quercetin and kaempferol, while SiRhaT68-1 was active toward isoquercetin and one of the four tested flavone 7-O-glucosides, diosmetin-7-O-glucoside. Kinetic analysis revealed an apparent difference in catalytic efficiency between the two enzymes under the conditions tested (SiUGT03: Km = 36.08 μM, kcat/Km = 0.01983 μM−1·s−1; SiRhaT68-1: Km = 184.70 μM, kcat/Km = 0.0005755 μM−1·s−1); however, since the Km of SiRhaT68-1 was close to the highest substrate concentration tested, this difference should be regarded as preliminary. These results, obtained from separate in vitro assays, demonstrated that the two enzymes catalyze two consecutive terminal glycosylation steps of rutin biosynthesis and represent promising candidates for further exploration of rutin biosynthesis in S. involucrata.
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BibTeX
@article{yi2026functional,
title = {{Functional Characterization of Glycosyltransferases from Saussurea involucrata Involved in Rutin Biosynthesis}},
author = {Peiqin Yi and Bingying Huang and Ruoting Zhan and Dongming Ma},
journal = {International Journal of Molecular Sciences},
year = {2026},
month = oct,
doi = {10.3390/ijms27219660},
url = {https://www.mdpi.com/1422-0067/27/21/9660},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Functional Characterization of Glycosyltransferases from Saussurea involucrata Involved in Rutin Biosynthesis
AU - Peiqin Yi
AU - Bingying Huang
AU - Ruoting Zhan
AU - Dongming Ma
PY - 2026
DA - 2026/10/28
JO - International Journal of Molecular Sciences
DO - 10.3390/ijms27219660
UR - https://www.mdpi.com/1422-0067/27/21/9660
AB - Saussurea involucrata is a medicinal plant rich in flavonoids, but the glycosyltransferases (GTs) responsible for flavonoid glycosylation in this species remain largely uncharacterized. Here, five GT genes (SiUGT03, SiUGT09, SiUGT60, SiRhaT68-1, and SiRhaT68-2) were identified. Four of the five proteins (SiUGT03, SiUGT09, SiUGT60, and SiRhaT68-1) possessed an intact PSPG-box, and phylogenetic analysis placed SiUGT03 in the 3GT clade and SiRhaT68-1 in the 1,6GT clade. Recombinant proteins were expressed in Escherichia coli as soluble maltose-binding protein (MBP) fusions and purified. In vitro enzymatic assays showed that SiUGT03 catalyzed the 3-O-glucosylation of quercetin to isoquercetin (quercetin-3-O-glucoside), whereas SiRhaT68-1 catalyzed the 1,6-rhamnosylation of isoquercetin to rutin. Among the substrates tested, SiUGT03 was active only toward the flavonols quercetin and kaempferol, while SiRhaT68-1 was active toward isoquercetin and one of the four tested flavone 7-O-glucosides, diosmetin-7-O-glucoside. Kinetic analysis revealed an apparent difference in catalytic efficiency between the two enzymes under the conditions tested (SiUGT03: Km = 36.08 μM, kcat/Km = 0.01983 μM−1·s−1; SiRhaT68-1: Km = 184.70 μM, kcat/Km = 0.0005755 μM−1·s−1); however, since the Km of SiRhaT68-1 was close to the highest substrate concentration tested, this difference should be regarded as preliminary. These results, obtained from separate in vitro assays, demonstrated that the two enzymes catalyze two consecutive terminal glycosylation steps of rutin biosynthesis and represent promising candidates for further exploration of rutin biosynthesis in S. involucrata.
SN - 1422-0067
ER -